Peripheral antinociceptive effects of a bifunctional µ and δ opioid receptor ligand in rat model of inflammatory bladder pain.

There is a need to develop a novel analgesic for pain associated with interstitial cystitis/painful bladder syndrome (IC/PBS). The use of the conventional µ-opioid receptor agonists to manage IC/PBS pain is controversial due to adverse CNS effects. These effects are attenuated in benzylideneoxymorphone (BOM), a low-efficacy µ-opioid receptor agonist/δ-opioid receptor antagonist that attenuates thermal pain and is devoid of reinforcing effects. We hypothesize that BOM will inhibit bladder pain by attenuating responses of urinary bladder distension (UBD)-sensitive afferent fibers. Therefore, the effect of BOM was tested on responses of UBD-sensitive afferent fibers in L6 dorsal root from inflamed and non-inflamed bladder of rats. Immunohistochemical (IHC) examination reveals that following the induction of inflammation there were significant high expressions of µ, δ, and µ-δ heteromer receptors in DRG. BOM dose-dependently (1-10 mg/kg, i.v) attenuated mechanotransduction properties of these afferent fibers from inflamed but not from non-inflamed rats. In behavioral model of bladder pain, BOM significantly attenuated visceromotor responses (VMRs) to UBD only in inflamed group of rats when injected either systemically (10 mg/kg, i.v.) or locally into the bladder (0.1 ml of 10 mg/ml). Furthermore, oxymorphone (OXM), a high-efficacy µ-opioid receptor agonist, attenuated responses of mechanosensitive bladder afferent fibers and VMRs to UBD. Naloxone (10 mg/kg, i.v.) significantly reversed the inhibitory effects of BOM and OXM on responses of bladder afferent fibers and VMRs suggesting µ-opioid receptor-related analgesic effects of these compounds. The results reveal that a low-efficacy, bifunctional opioid-based compound can produce analgesia by attenuating mechanotransduction functions of afferent fibers innervating the urinary bladder.

Temporal gene expression changes after acute and delayed ventral root avulsion-reimplantation.

BACKGROUND: In a model of injured spinal motor neurons where the avulsed spinal nerve is surgically reimplanted, useful regrowth of the injured nerve follows, both in animal experiments and clinical cases. This has led to surgical reimplantation strategies with subsequent partial functional motoric recovery. Still, the ideal time point for successful regeneration after reimplantation and the specific genetic profile of this time point is not known. OBJECTIVE: To explore the temporal gene expression of the whole genome in the ventral spinal cord after reimplantation at different time points after avulsion. METHODS: Totally 18 adult rats were subjected to avulsion of the left L5 root only (N = 3), avulsion followed by acute spinal reimplantation (N = 3), avulsion followed by 24 h (N = 3) or 48 h (N = 3) delayed reimplantation. Animals were allowed to survive 24 h after their respective surgery whereafter the ventral quadrant of the spinal cord at the operated side was harvested, processed for and analysed with Affymetrix Rat Gene ST 1.0 array followed by statistical analysis of gene expression patternsResults:Specific gene expression patterns were found at different time points after avulsion and reimplantation. Over all, early reimplantation seemed to diminish inflammatory response and support gene regulation related to neuronal activity compared to avulsion only or delayed reimplantation. In addition did gene activity after avulsion-reimplantation correspond to regeneration-associated genes typical for regeneration in the peripheral nervous system. CONCLUSIONS: Our study reveal that genetic profiling after this kind of injury is possible, that specific and distinct expression patterns can be found with early reimplantation being favourable over late and that regenerative activity in this kind of injury bears hallmark typical for peripheral nerve regeneration. These findings can be useful in elucidating specific genetic expression typical for successful nerve regeneration, hopefully not only in this specific model but in the nervous system in general.

Neuropeptide Y release in the rat spinal cord measured with Y1 receptor internalization is increased after nerve injury.

Neuropeptide Y (NPY) modulates nociception in the spinal cord, but little is known about its mechanisms of release. We measured NPY release in situ using the internalization of its Y1 receptor in dorsal horn neurons. Y1 receptor immunoreactivity was normally localized to the cell surface, but addition of NPY to spinal cord slices increased the number of neurons with Y1 internalization in a biphasic fashion (EC50s of 1 nM and 1 µM). Depolarization with KCl, capsaicin, or the protein kinase A activator 6-benzoyl-cAMP also induced Y1 receptor internalization, presumably by releasing NPY. NMDA receptor activation in the presence of BVT948, an inhibitor of protein tyrosine phosphatases, also released NPY. Electrical stimulation of the dorsal horn frequency-dependently induced NPY release; and this was decreased by the Y1 antagonist BIBO3304, the Nav channel blocker lidocaine, or the Cav2 channel blocker ω-conotoxin MVIIC. Dorsal root immersion in capsaicin, but not its electrical stimulation, also induced NPY release. This was blocked by CNQX, suggesting that part of the NPY released by capsaicin was from dorsal horn neurons receiving synapses from primary afferents and not from the afferent themselves. Mechanical stimulation in vivo, with rub or clamp of the hindpaw, elicited robust Y1 receptor internalization in rats with spared nerve injury but not sham surgery. In summary, NPY is released from dorsal horn interneurons or primary afferent terminals by electrical stimulation and by activation of TRPV1, PKA or NMDA receptors in. Furthermore, NPY release evoked by noxious and tactile stimuli increases after peripheral nerve injury.

AMP-Activated Protein Kinase Activation in Dorsal Root Ganglion Suppresses mTOR/p70S6K Signaling and Alleviates Painful Radiculopathies in Lumbar Disc Herniation Rat Model.

STUDY DESIGN: Animal experiment: a rat model of lumbar disc herniation (LDH) induced painful radiculopathies. OBJECTIVE: To investigate the role and mechanism of AMP-activated protein kinase (AMPK) in dorsal root ganglia (DRG) neurons in LDH-induced painful radiculopathies. SUMMARY OF BACKGROUND DATA: Overactivation of multiple pain signals in DRG neurons triggered by LDH is crucial to the development of radicular pain. AMPK is recognized as a cellular energy sensor, as well as a pain sensation modulator, but its function in LDH-induced pain hypersensitivity remains largely unknown. METHODS: The LDH rat model was established by autologous nucleus pulposus transplantation into the right lumbar 5 (L5) nerve root. At different time points after AMPK agonist metformin (250 mg/kg/d) or mammalian target of rapamycin (mTOR) inhibitor rapamycin (5 mg/kg) intraperitoneal administration, thermal and mechanical sensitivity were evaluated by measuring paw withdrawal latency (PWL) and 50% paw withdrawal thresholds (PWT). The levels of AMPK, mTOR, and p70S6K phosphorylation were determined by Western blot. We also investigated the proportion of p-AMPK positive neurons in the right L5 DRG neurons using immunofluorescence. RESULTS: LDH evoked persistent thermal hyperalgesia and mechanical allodynia on the ipsilateral paw, as indicated by the decreased PWL and 50% PWT. These pain hypersensitive behaviors were accompanied with significant inhibition of AMPK and activation of mTOR in the associated DRG neurons. Pharmacological activation of AMPK in the DRG neurons not only suppressed mTOR/p70S6K signaling, but also alleviated LDH-induced pain hypersensitive behaviors. CONCLUSION: We provide a molecular mechanism for the activation of pain signals based on AMPK-mTOR axis, as well as an intervention strategy by targeting AMPK-mTOR axis in LDH-induced painful radiculopathies. LEVEL OF EVIDENCE: N/A.

Microglia exit the CNS in spinal root avulsion.

Microglia are central nervous system (CNS)-resident cells. Their ability to migrate outside of the CNS, however, is not understood. Using time-lapse imaging in an obstetrical brachial plexus injury (OBPI) model, we show that microglia squeeze through the spinal boundary and emigrate to peripheral spinal roots. Although both macrophages and microglia respond, microglia are the debris-clearing cell. Once outside the CNS, microglia re-enter the spinal cord in an altered state. These peripheral nervous system (PNS)-experienced microglia can travel to distal CNS areas from the injury site, including the brain, with debris. This emigration is balanced by two mechanisms-induced emigration via N-methyl-D-aspartate receptor (NMDA) dependence and restriction via contact-dependent cellular repulsion with macrophages. These discoveries open the possibility that microglia can migrate outside of their textbook-defined regions in disease states.

Possible involvement of peripheral TRP channels in the hydrogen sulfide-induced hyperalgesia in diabetic rats.

BACKGROUND: Peripheral diabetic neuropathy can be painful and its symptoms include hyperalgesia, allodynia and spontaneous pain. Hydrogen sulfide (H2S) is involved in diabetes-induced hyperalgesia and allodynia. However, the molecular target through which H2S induces hyperalgesia in diabetic animals is unclear. The aim of this study was to determine the possible involvement of transient receptor potential (TRP) channels in H2S-induced hyperalgesia in diabetic rats. RESULTS: Streptozotocin (STZ) injection produced hyperglycemia in rats. Intraplantar injection of NaHS (an exogenous donor of H2S, 3-100 µg/paw) induced hyperalgesia, in a time-dependent manner, in formalin-treated diabetic rats. NaHS-induced hyperalgesia was partially prevented by local intraplantar injection of capsazepine (0.3-3 µg/paw), HC-030031 (100-316 µg/paw) and SKF-96365 (10-30 µg/paw) blockers, at 21 days post-STZ injection. At the doses used, these blockers did not modify formalin-induced nociception. Moreover, capsazepine (0.3-30 µg/paw), HC-030031 (100-1000 µg/paw) and SKF-96365 (10-100 µg/paw) reduced formalin-induced nociception in diabetic rats. Contralateral injection of the highest doses used did not modify formalin-induced flinching behavior. Hyperglycemia, at 21 days, also increased protein expression of cystathionine-ß-synthase enzyme (CBS) and TRPC6, but not TRPA1 nor TRPV1, channels in dorsal root ganglia (DRG). Repeated injection of NaHS enhanced CBS and TRPC6 expression, but hydroxylamine (HA) prevented the STZ-induced increase of CBS protein. In addition, daily administration of SKF-96365 diminished TRPC6 protein expression, whereas NaHS partially prevented the decrease of SKF-96365-induced TRPC6 expression. Concordantly, daily intraplantar injection of NaHS enhanced, and HA prevented STZ-induced intraepidermal fiber loss, respectively. CBS was expressed in small- and medium-sized cells of DRG and co-localized with TRPV1, TRPA1 and TRPC6 in IB4-positive neurons. CONCLUSIONS: Our data suggest that H2S leads to hyperalgesia in diabetic rats through activation of TRPV1, TRPA1 and TRPC channels and, subsequent intraepidermal fibers loss. CBS enzyme inhibitors or TRP-channel blockers could be useful for treatment of painful diabetic neuropathy.

trans-Anethole of Fennel Oil is a Selective and Nonelectrophilic Agonist of the TRPA1 Ion Channel.

Transient receptor potential (TRP) cation channels are molecular targets of various natural products. TRPA1, a member of TRP channel family, is specifically activated by natural products such as allyl isothiocyanate (mustard oil), cinnamaldehyde (cinnamon), and allicin (garlic). In this study, we demonstrated that TRPA1 is also a target of trans-anethole in fennel oil (FO) and fennel seed extract. Similar to FO, trans-anethole selectively elicited calcium influx in TRPA1-expressing mouse sensory neurons of the dorsal root and trigeminal ganglia. These FO- and anethole-induced calcium responses were blocked by a selective TRPA1 channel antagonist, HC-030031. Moreover, both FO and trans-anethole induced calcium influx and transmembrane currents in HEK293 cells stably overexpressing human TRPA1 channels, but not in regular HEK293 cells. Mutation of the amino acids S873 and T874 binding site of human TRPA1 significantly attenuated channel activation by trans-anethole, whereas pretreating with glutathione, a nucleophile, did not. Conversely, activation of TRPA1 by the electrophile allyl isothiocyanate was abolished by glutathione, but was ostensibly unaffected by mutation of the ST binding site. Finally, it was found that trans-anethole was capable of desensitizing TRPA1, and unlike allyl isothiocyanate, it failed to induce nocifensive behaviors in mice. We conclude that trans-anethole is a selective, nonelectrophilic, and seemingly less-irritating agonist of TRPA1.

Alterations in the spinal cord and ventral root after cerebral infarction in non-human primates.

BACKGROUNDS: Cerebral infarction does not only cause focal injury in the ischemic site, but also secondary non-ischemic damage at the remote areas of nervous system associated with the primary focus. OBJECTIVE: This study investigated the changes in the spinal cord and ventral root after middle cerebral artery occlusion (MCAO) in cynomolgus monkeys (Macaca fascicularis). METHODS: Adult male cynonolgus monkeys (4-5 years, 5.5-7.5 kg) were subjected to MCAO (n = 6) or sham surgery (n = 4). After 12 weeks, spinal cords and the ventral roots were harvested. Morphometric alterations in the spinal cord were detected at C5 and L5 levels via immunofluorescence. The profiles of C5 and L5 ventral roots were displayed by toluidine blue staining and transmission electron microscopic examination. RESULTS: Significant axonal loss in the contralateral corticospinal tract and abnormally enlarged axons in the ipsilateral were observed in monkeys with MCAO. The number of neurons in the contralateral ventral horn got declined while that in the ipsilateral was almost unaffected after MCAO compared with sham controls. Glial activation post-MCAO was observed in the bilateral corticospinal tract and the ventral horn. Aberrant nerve fibers appeared frequently in the contralateral ventral roots of MCAO monkey but rarely in the ipsilateral. CONCLUSIONS: These results indicate that focal cerebral infarction leads to pathological alterations in the spinal cord and ventral roots in non-human primates.

Iron-specific Signal Separation from within Heavy Metal Stained Biological Samples Using X-Ray Microtomography with Polychromatic Source and Energy-Integrating Detectors.

Biological samples are frequently stained with heavy metals in preparation for examining the macro, micro and ultra-structure using X-ray microtomography and electron microscopy. A single X-ray microtomography scan reveals detailed 3D structure based on staining density, yet it lacks both material composition and functional information. Using a commercially available polychromatic X-ray source, energy integrating detectors and a two-scan configuration labelled by their energy- "High" and "Low", we demonstrate how a specific element, here shown with iron, can be detected from a mixture with other heavy metals. With proper selection of scan configuration, achieving strong overlap of source characteristic emission lines and iron K-edge absorption, iron absorption was enhanced enabling K-edge imaging. Specifically, iron images were obtained by scatter plot material analysis, after selecting specific regions within scatter plots generated from the "High" and "Low" scans. Using this method, we identified iron rich regions associated with an iron staining reaction that marks the nodes of Ranvier along nerve axons within mouse spinal roots, also stained with osmium metal commonly used for electron microscopy.

Netrin-1 Confines Rhombic Lip-Derived Neurons to the CNS.

During brainstem development, newborn neurons originating from the rhombic lip embark on exceptionally long migrations to generate nuclei important for audition, movement, and respiration. Along the way, this highly motile population passes several cranial nerves yet remains confined to the CNS. We found that Ntn1 accumulates beneath the pial surface separating the CNS from the PNS, with gaps at nerve entry sites. In mice null for Ntn1 or its receptor DCC, hindbrain neurons enter cranial nerves and migrate into the periphery. CNS neurons also escape when Ntn1 is selectively lost from the sub-pial region (SPR), and conversely, expression of Ntn1 throughout the mutant hindbrain can prevent their departure. These findings identify a permissive role for Ntn1 in maintaining the CNS-PNS boundary. We propose that Ntn1 confines rhombic lip-derived neurons by providing a preferred substrate for tangentially migrating neurons in the SPR, preventing their entry into nerve roots.

Early CALP2 expression and microglial activation are potential inducers of spinal IL-6 up-regulation and bilateral pain following motor nerve injury.

Previous work from our laboratory showed that motor nerve injury by lumbar 5 ventral root transection (L5-VRT) led to interleukin-6 (IL-6) over-expression in bilateral spinal cord, and that intrathecal administration of IL-6 neutralizing antibody delayed the induction of mechanical allodynia in bilateral hind paws. However, early events and upstream mechanisms underlying spinal IL-6 expression following L5-VRT require elucidation. The model of L5-VRT was used to induce neuropathic pain, which was assessed with von Frey hairs and the plantar tester in adult male Sprague-Dawley rats. Calpain-2 (CALP2, a calcium-dependent protease) knockdown or over-expression and microglia depletion were conducted intrathecally. Western blots and immunohistochemistry were performed to explore the possible mechanisms. Here, we provide the first evidence that both IL-6 and CALP2 levels are increased in lumbar spinal cord within 30 min following L5-VRT. IL-6 and CALP2 co-localized in both spinal dorsal horn (SDH) and spinal ventral horn. Post-operative (PO) increase in CALP2 in ipsilateral SDH was evident at 10 min PO, preceding increased IL-6 at 20 min PO. Knockdown of spinal CALP2 by intrathecal CALP2-shRNA administration prevented VRT-induced IL-6 overproduction in ipsilateral spinal cord and alleviated bilateral mechanical allodynia. Spinal microglia activation also played a role in early IL-6 up-regulation. Macrophage/microglia markers ED1/Iba1 were increased at 30 min PO, while glial fibrillary acidic protein (astrocyte) and CNPase (oligodendrocyte) markers were not. Increased Iba1 was detected as early as 20 min PO and peaked at 3 days. Morphology changed from a small soma with fine processes in resting cells to an activated ameboid shape. Depletion of microglia using Mac-1-saporin partially prevented IL-6 up-regulation and attenuated VRT-induced bilateral mechanical allodynia. Taken together, our findings provide evidence that increased spinal cord CALP2 and microglia cell activation may have early causative roles in IL-6 over-expression following motor nerve injury. Agents that inhibit CALP2 and/or microglia activation may therefore prove valuable for treating neuropathic pain.

Calpain-2 Regulates TNF-α Expression Associated with Neuropathic Pain Following Motor Nerve Injury.

Both calpain-2 (CALP2) and tumor necrosis factor-α (TNF-α) contribute to persistent bilateral hypersensitivity in animals subjected to L5 ventral root transection (L5-VRT), a model of selective motor fiber injury without sensory nerve damage. However, specific upstream mechanisms regulating TNF-α overexpression and possible relationships linking CALP2 and TNF-α have not yet been investigated in this model. We examined changes in CALP2 and TNF-α protein levels and alterations in bilateral mechanical threshold within 24 h following L5-VRT model injury. We observed robust elevation of CALP2 and TNF-α in bilateral dorsal root ganglias (DRGs) and bilateral spinal cord neurons. CALP2 and TNF-α protein induction by L5-VRT were significantly inhibited by pretreatment using the calpain inhibitor MDL28170. Administration of CALP2 to rats without nerve injury further supported a role of CALP2 in the regulation of TNF-α expression. Although clinical trials of calpain inhibition therapy for alleviation of neuropathic pain induced by motor nerve injury have not yet shown success, our observations linking CALP2 and TNF-α provide a framework of a systems' approach based perspective for treating neuropathic pain.

TNFa/TNFR2 signaling is required for glial ensheathment at the dorsal root entry zone.

Somatosensory information from the periphery is routed to the spinal cord through centrally-projecting sensory axons that cross into the central nervous system (CNS) via the dorsal root entry zone (DREZ). The glial cells that ensheath these axons ensure rapid propagation of this information. Despite the importance of this glial-axon arrangement, how this afferent nerve is assembled during development is unknown. Using in vivo, time-lapse imaging we show that as centrally-projecting pioneer axons from dorsal root ganglia (DRG) enter the spinal cord, they initiate expression of the cytokine TNFalpha. This induction coincides with ensheathment of these axons by associated glia via a TNF receptor 2 (TNFR2)-mediated process. This work identifies a signaling cascade that mediates peripheral glial-axon interactions and it functions to ensure that DRG afferent projections are ensheathed after pioneer axons complete their navigation, which promotes efficient somatosensory neural function.

Melatonin prevents mitochondrial dysfunction and promotes neuroprotection by inducing autophagy during oxaliplatin-evoked peripheral neuropathy.

Oxaliplatin, an organoplatinum compound, is used in the treatment of colorectal cancer, but its clinical use can be limited due to the development of peripheral neuropathy. Whilst mitochondrial dysfunction has been implicated as a major pathomechanism for oxaliplatin-induced neurotoxicity, the prevention of autophagy may also aggravate neuronal cell death. Melatonin, a well-known mitoprotectant and autophagy inducer, was used to examine its neuroprotective role in oxaliplatin-induced peripheral neuropathy (OIPN). Melatonin prevented the loss of mitochondrial membrane potential (Ψm) and promoted neuritogenesis in oxaliplatin-challenged neuro-2a cells. It did not interfere with the cytotoxic activity of oxaliplatin in human colon cancer cell line, HT-29. Melatonin treatment significantly alleviated oxaliplatin-induced pain behavior and neuropathic deficits in rats. It also ameliorated nitro-oxidative stress mediated by oxaliplatin, thus prevented nitrosylation of proteins and loss of antioxidant enzymes, and therefore, it improved mitochondrial electron transport chain function and maintained cellular bioenergetics by improving the ATP levels. The protective effects of melatonin were attributed to preventing oxaliplatin-induced neuronal apoptosis by increasing the autophagy pathway (via LC3A/3B) in peripheral nerves and dorsal root ganglion (DRG). Hence, it preserved the epidermal nerve fiber density in oxaliplatin-induced neuropathic rats. Taken together, we provide detailed molecular mechanisms for the neuroprotective effect of melatonin and suggest it has translational potential for oxaliplatin-induced neuropathy.

A Morphological and Molecular Characterization of the Spinal Cord after Ventral Root Avulsion or Distal Peripheral Nerve Axotomy Injuries in Adult Rats.

Retrograde cell death in sensory dorsal root ganglion cells following peripheral nerve injury is well established. However, available data regarding the underlying mechanism behind injury induced motoneuron death are conflicting. By comparing morphological and molecular changes in spinal motoneurons after L4-L5 ventral root avulsion (VRA) and distal peripheral nerve axotomy (PNA) 7 and 14 days postoperatively, we aimed to gain more insight about the mechanism behind injury-induced motoneuron degeneration. Morphological changes in spinal cord were assessed by using quantitative immunohistochemistry. Neuronal degeneration was revealed by decreased immunostaining for microtubule-associated protein-2 in dendrites and synaptophysin in presynaptic boutons after both VRA and PNA. Significant motoneuron atrophy was already observed at 7 days post-injury, independently of injury type. Immunostaining for ED1 reactive microglia was significantly elevated in all experimental groups, as well as the astroglial marker glial fibrillary acidic protein (GFAP). Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis of the ventral horn from L4-L5 spinal cord segments revealed a significant upregulation of genes involved in programmed cell death including caspase-3, caspase-8, and related death receptors TRAIL-R, tumor necrosis factor (TNF)-R, and Fas following VRA. In contrast, following PNA, caspase-3 and the death receptor gene expression levels did not differ from the control, and there was only a modest increased expression of caspase-8. Moreover, the altered gene expression correlated with protein changes. These results show that the spinal motoneurons reacted in a similar fashion with respect to morphological changes after both proximal and distal injury. However, the increased expression of caspase-3, caspase-8, and related death receptors after VRA suggest that injury- induced motoneuron degeneration is mediated through an apoptotic mechanism, which might involve both the intrinsic and the extrinsic pathways.

Direct Spinal Ventral Root Repair following Avulsion: Effectiveness of a New Heterologous Fibrin Sealant on Motoneuron Survival and Regeneration.

Axonal injuries at the interface between central and peripheral nervous system, such as ventral root avulsion (VRA), induce important degenerative processes, mostly resulting in neuronal and motor function loss. In the present work, we have compared two different fibrin sealants, one derived from human blood and another derived from animal blood and Crotalus durissus terrificus venom, as a promising treatment for this type of injury. Lewis rats were submitted to VRA (L4-L6) and had the avulsed roots reimplanted to the surface of the spinal cord, with the aid of fibrin sealant. The spinal cords were processed to evaluate neuronal survival, synaptic stability, and glial reactivity, 4 and 12 weeks after lesion. Sciatic nerves were processed to investigate Schwann cell activity by p75(NTR) expression (4 weeks after surgery) and to count myelinated axons and morphometric evaluation (12 weeks after surgery). Walking track test was used to evaluate gait recovery, up to 12 weeks. The results indicate that both fibrin sealants are similarly efficient. However, the snake-derived fibrin glue is a potentially safer alternative for being a biological and biodegradable product which does not contain human blood derivatives. Therefore, the venom glue can be a useful tool for the scientific community due to its advantages and variety of applications.

Expressions of miR-132, miR-134, and miR-485 in rat primary motor cortex during transhemispheric functional reorganization after contralateral seventh cervical spinal nerve root transfer following brachial plexus avulsion injuries.

The transfer of a contralateral healthy seventh cervical spinal nerve root (cC7) to the recipient nerve in the injured side is considered a promising procedure for restoration of the physiological functions of an injured hand after brachial plexus root avulsion injury (BPAI). Growing evidence shows that transhemispheric cortical reorganization plays an important role in the functional recovery of the injured arm after cC7 nerve transfer surgery. However, the molecular mechanism underlying the transhemispheric cortical reorganization after cC7 transfer remains elusive. In the present study, we investigated the expression of miR-132, miR-134, and miR-485 in the rat primary motor cortex after cC7 transfer following BPAI by quantitative PCR. The results demonstrated the dynamic alteration in the expression of miR-132, miR-134, and miR-485 in the primary motor cortex of rats after cC7 transfer following BPAI. It indicates that microRNAs are involved in the dynamic transhemispheric functional reorganization after cC7 root transfer following BPAI. Together, this study is the first to provide evidence for the involvement of microRNAs during dynamic transhemispheric functional reorganization after cC7 transfer following BPAI. The results are useful for understanding the mechanism underlying transhemispheric functional reorganization after contralateral seventh cervical spinal nerve root transfer following BPAI.

Clinical and neurobiological advances in promoting regeneration of the ventral root avulsion lesion.

Root avulsions due to traction to the brachial plexus causes complete and permanent loss of function. Until fairly recent, such lesions were considered impossible to repair. Here we review clinical repair strategies and current progress in experimental ventral root avulsion lesions. The current gold standard in patients with a root avulsion is nerve transfer, whereas reimplantation of the avulsed root into the spinal cord has been performed in a limited number of cases. These neurosurgical repair strategies have significant benefit for the patient but functional recovery remains incomplete. Developing new ways to improve the functional outcome of neurosurgical repair is therefore essential. In the laboratory, the molecular and cellular changes following ventral root avulsion and the efficacy of intervention strategies have been studied at the level of spinal motoneurons, the ventral spinal root and peripheral nerve, and the skeletal muscle. We present an overview of cell-based pharmacological and neurotrophic factor treatment approaches that have been applied in combination with surgical reimplantation. These interventions all demonstrate neuroprotective effects on avulsed motoneurons, often accompanied with various degrees of axonal regeneration. However, effects on survival are usually transient and robust axon regeneration over long distances has as yet not been achieved. Key future areas of research include finding ways to further extend the post-lesion survival period of motoneurons, the identification of neuron-intrinsic factors which can promote persistent and long-distance axon regeneration, and finally prolonging the pro-regenerative state of Schwann cells in the distal nerve.

Overexpression of ATF3 or the combination of ATF3, c-Jun, STAT3 and Smad1 promotes regeneration of the central axon branch of sensory neurons but without synergistic effects.

Peripheral nerve injury results in the activation of a number of transcription factors (TFs) in injured neurons, some of which may be key regulators of the regeneration-associated gene (RAG) programme. Among known RAG TFs, ATF3, Smad1, STAT3 and c-Jun have all been linked to successful axonal regeneration and have known functional and physical interactions. We hypothesised that TF expression would promote regeneration of the central axon branch of DRG neurons in the absence of a peripheral nerve lesion and that simultaneous overexpression of multiple RAG TFs would lead to greater effects than delivery of a single TF. Using adeno-associated viral vectors, we overexpressed either the combination of ATF3, Smad1, STAT3 and c-Jun with farnesylated GFP (fGFP), ATF3 only with fGFP, or fGFP only, in DRG neurons and assessed axonal regeneration after dorsal root transection or dorsal column injury and functional improvement after dorsal root injury. ATF3 alone and the combination of TFs promoted faster regeneration in the injured dorsal root. Surprisingly, however, the combination did not perform better than ATF3 alone. Neither treatment was able to induce functional improvement on sensory tests after dorsal root injury or promote regeneration in a dorsal column injury model. The lack of synergistic effects among these factors indicates that while they do increase the speed of axon growth, there may be functional redundancy between these TFs. Because axon growth is considerably less than that seen after a conditioning lesion, it appears these TFs do not induce the full regeneration programme.